Search

BackgroundThis report presents a case of an adult Polish women of Caucasian origin who was heterozygous for the nondeletional mutation: Hb Handsworth (HBA2 or HBA1: c.55G > C, p.Gly19Arg) and deletional (-α3.7) α-thalassemia mutation.MethodsThe HbA2 and HbF levels were measured by microcolumn chromatography and alkaline denaturation procedure, respectively, while electrophoresis was used to detect pathological hemoglobin fraction. The β- and α-globin genotypes were determined by DNA sequencing, gap-polymerase chain reaction, α gene triplication and MLPA.ResultsThe HbA2 and HbF levels were normal, but hemoglobin electrophoresis on agarose gel alkaline pH showed a strong band migration in a position of hemoglobin S and faint bands in the neighborhood of band A on acid electrophoresis. Molecular analysis of the alpha globin cluster detected a point mutation at codon 19 in HBA2 (c.55G > C, p.Gl- y19Arg) and deletion -α3.7.ConclusionsOur compound heterozygosity does not produce severe clinical or hematological symptoms but it is important to say that in our part of Europe such cases do appear. Molecular analysis of the alpha globin cluster is required for correct diagnosis in patients with normal HbA2 levels. Compound heterozygosity was unmasked by molecular diagnosis only.

Mutations in isocitrate dehydrogenase 1 and 2 (IDH1/2) genes occur in about 20% patients with acute myeloid leukemia (AML), leading to DNA hypermethylation and epigenetic deregulation. We assessed the prognostic significance of IDH1/2 mutations (IDH1/2+) in 398 AML patients with normal karyotype (NK-AML), treated with daunorubicine + cytarabine (DA), DA + cladribine (DAC), or DA + fludarabine. IDH2 mutation was an independent favorable prognostic factor for 4-year overall survival (OS) in total NK-AML population (p = 0.03, censoring at allotransplant). We next evaluated the effect of addition of cladribine to induction regimen on the patients’ outcome according to IDH1/2 mutation status. In DAC group, 4-year OS was increased in IDH2+ patients, compared to IDH-wild type group (54% vs 33%; p = 0.0087, censoring at allotransplant), while no difference was observed for DA-treated subjects. In multivariate analysis, DAC independently improved the survival of IDH2+ patients (HR = 0.6 [0.37–0.93]; p = 0.024; censored at transplant), indicating that this group specifically benefits from cladribine-containing therapy. In AML cells with R140Q or R172K IDH2 mutations, cladribine restrained mutations-related DNA hypermethylation. Altogether, DAC regimen produces better outcomes in IDH2+ NK-AML patients than DA, and this likely results from the hypomethylating activity of cladribine. Our observations warrant further investigations of induction protocols combining cladribine with IDH1/2 inhibitors in IDH2-mutant.

Autologous hematopoietic stem cell transplantation (autoHSCT) requires collection of sufficient number of hematopoietic stem cells. The goal of this study was to evaluate efficacy of plerixafor used in patients with lymphoid malignancies failing conventional stem cell mobilization.This was a prospective, non-interventional study. All consecutive patients (n = 109) treated with plerixafor in 11 centers were reported. The drug was used either in case of previous mobilization failure (n = 67) or interventionally, in case of insufficient CD34+ cell output during current mobilization (n = 42). Successful mobilization was defined as resulting in collection of ≥ 2 × 106 CD34+ cells/kg for single autoHSCT or ≥ 4 × 106 CD34+ cells/kg for double procedure.The overall rate of successful mobilization was 55% (55% for single and 56% for double autoHSCT). The median total number of collected CD34+ cells/kg was 2.4 (range, 0-11.5) for patients intended for a single transplantation while 4.0 (0.6-16.9) for double procedure. The number of circulating CD34+ cells increased after the use of plerixafor regardless of baseline values. The median fold increase was 3.3 (0.3-155). Data from this observational study confirm high efficacy of plerixafor used in routine clinical practice as salvage for patients with lymphoid malignancies failing conventional stem cell mobilization.

In 23 patients with polycythaemia vera and symptomatic erythrocytosis, glomerular filtration rate and urine concentration ability see article were determined under conditions of antidiuresis. The restriction of fluid intake caused a significant reduction of GFR and modified the osmotic function of the kidneys. The results were similar in both types of polycythaemia.

The long term prognosis of adult patients with relapsed/refractory (R/R) acute lymphoblastic leu­kemia (ALL) is poor. There is no standard treatment for relapsed ALL and reinduction therapy should only be a “bridge treatment” in allogeneic hematopoietic stem cell transplantation (allo- -HSCT). In spite of obtaining an initial response to treatment, the duration of response using standard chemotherapy is usually short and the next relapses are chemoresistant. Immunotherapy with monoclonal antibodies as a targeted therapy represents a novel approach for treating patients with R/R ALL. The objective of immunotherapy is to improve the relapse-free and overall survival without increasing the toxicity of treatment. In this review, the mechanism of action and clini­cal efficacy of the novel bispecific antibody blinatumomab is discussed. The European Medicines Agency approved blinatumomab in 2015 for treating adult patients with Philadelphia chromosome negative R/R B-ALL.

Background Alpha-thalassemia is genetically transmitted hemolytic anemia resulting from disturbance of the globins chain synthesis. Alpha-thalassemia is caused most frequently by deletions and less commonly by nondeletional defects. Aim To introduce the molecular methods for routine identifications of alpha-thalassemia mutations and to study the characteristics of these mutations in Poland. Methods Blood sample of 155 patients with normal or reduced HbA2 values was obtained for blood counting. All samples underwent gap-PCR to screen for the seven common α-thal deletions and MLPA analysis. The DNA of 21 patients in which deletions were not detected has been directly sequenced. Results We detected mutations in the alpha-globin gene in 72 of 155 patients studied. 55 out of 72 cases showed most common thalassemia deletions, as the following: a single gene deletion (α3.7 and α4.2) and both genes deletion (FIL, SEA, MED I, and α20.5). Owing to the use of MLPA technique, we found nontypical deletions in another 12 patients and multiplication of the alpha-globin genes in further 4 cases. We also identified a patient with a point mutation HBA2: c.300 + 2T>A by DNA sequencing. Conclusions Molecular analysis of the alpha-globin cluster is required for a correct diagnosis in patients with normal or reduced level of HbA2. The results of the study show that due to the progressive migration of the population and globalization, thalassemia must be included in the differential diagnosis of anemia in Poland.

Monosomal karyotype (MK) and complex karyotype (CK) are poor prognostic factors in acute myeloid leukemia (AML). A comprehensive analysis of cytogenetic and clinical factors influencing an outcome of AML-CK+ was performed. The impact of cladribine containing induction on treatment results was also evaluated. We analyzed 125 patients with AML-CK+ treated within PALG protocols. MK was found in 75 (60%) individuals. The overall complete remission (CR) rate of 66 intensively treated patients was 62% vs. 28% in CK+ MK− and CK+ MK+ group (p = .01). No difference in CR rate was observed between DA and DAC arms. The overall survival (OS) in intensively treated patients was negatively influenced by MK, karyotype complexity (≥5 abnormalities), and WBC >20 G/L in multivariate analysis. The addition of cladribine to DA regimen improved OS only in MK− but not in MK+ group. In conclusion, concomitance of MK with ≥5 chromosomal abnormalities is associated with dismal treatment outcome in AMK-CK+.

Acquired copy number changes are common in acute leukemia. They are reported as recurrent amplifications or deletions (del), and may be indicative of involvement of oncogenes or tumor suppressor genes in acquired disease, as well as serving as potential biomarkers for prognosis or as targets for molecular therapy. The present study reported a gain of copy number of 14q13 to 14q32, leading to immunoglobulin heavy chain locus splitting in a young adult female. To the best of our knowledge, this rearrangement has not been previously reported in B-cell acute lymphoblastic leukemia (ALL). Low resolution banding cytogenetics performed at the time of diagnosis revealed a normal karyotype. However, retrospective application of fluorescence in situ hybridization (FISH) banding and locus-specific FISH probes, as well as multiplex ligation-dependent probe amplification and high resolution array-comparative genomic hybridization, revealed previously hidden aberrations. Overall, a karyotype of 46, XX, del(9) (p21.3 p21.3), derivative(14) (pter-> q32.33:: q32.33-> q13 ::q32.33-> qter) was determined. The patient was treated according to the Polish Adult Leukemia Group protocol and achieved complete remission. The results of the present study indicate that a favorable prognosis is associated with these aberrations when the aforementioned treatment is administered.

This prospective study estimated outcomes in 509 elderly patients with acute myeloid leukemia (AML) with different treatment approaches depending on Eastern Cooperative Oncology Group (ECOG) performance status and Charlson Comorbidity Index (CCI). Patients were stratified into fit (ECOG 0–2 and CCI 0–2) or frail (ECOG > 2 and/or CCI > 2) groups. Fit patients with CCI 0 received intensive chemotherapy whilst reduced-intensive chemotherapy (R-IC) was given to those with CCI 1–2. Frail patients received best supportive therapy. Fit patients presented a longer overall survival (OS) than frail subjects, but 8-week mortality rates were similar. The complete response (CR) rate between fit CCI 0 and CCI 1–2 subgroups was significantly different. Both of the fit subgroups showed similar 8-week mortality rates and OS probabilities. Allocating fit patients with CCI 1–2 to R-IC enabled an increase in the group of elderly patients who could be treated with the intention of inducing remission.

Acute acalculous cholecystitis (ACC) is most frequently reported in critically ill patients following sepsis, extensive injury or surgery. It is rather uncommon as a chemotherapy-induced complication, which is usually life-threatening in neutropenic patients subjected to myelosuppressive therapy. A 23-year-old patient with acute lymphoblastic leukemia was subjected to myelosuppressive chemotherapy (cyclophosphamide, cytarabine, pegaspargase). After the first chemotherapy cycle the patient was neutropenic and feverish; she presented with vomiting and pain in the right epigastrium. Ultrasound demonstrated an acalculous gallbladder with wall thickening up to 14 mm. The ACC was diagnosed. Medical therapy included a broad spectrum antibiotic regimen and granulocyte-colony stimulating factors. On the second day after ACC diagnosis the patient's general condition worsened. Laparoscopic cholecystectomy was performed. The resected gallbladder showed no signs of bacterial or leukemic infiltrates. The postoperative course was uneventful. In the management of neutropenic patients with ACC surgical treatment is as important as pharmacological therapy.

A monosomal karyotype (MK) was identified by banding techniques (BT) in acute myeloid leukemia (AML). However, BT may be insufficient to determine the actual loss of a complete chromosome, especially in complex karyotypes. We have investigated the effect of monosomy type, total (MK-t) and partial (MK-p), reevaluated by FISH, on prognosis. We have found that complete remission rate and probability of overall survival at 1 year was higher in MK-p (n=27) than MK-t (n=15) group (40% vs. 15.4%, P=0.19 and 30% vs. 9%, P=0.046, respectively). Our results indicate for the first time that monosomy type influences the prognosis of MK-AML.

To the editor: Internal tandem duplication in the FLT3 gene ( FLT3- ITD) has been recognized as a marker conferring poor outcome in patients with normal karyotype acute myeloid leukemia (NK-AML).[1][1] Because of the inferior outcome of FLT3- ITD+ NK-AML patients when treated with standard

Acute myeloid leukemia (AML) cells harbor frequent mutations in genes responsible for epigenetic modifications. Increasing evidence of clinical role of DNMT3A and IDH1/2 mutations highlights the need for a robust and inexpensive test to identify these mutations in routine diagnostic work-up. Herein, we compared routinely used direct sequencing method with high-resolution melting (HRM) assay for screening DNMT3A and IDH1/2 mutations in patients with AML. We show very high concordance between HRM and Sanger sequencing (100% samples for IDH2-R140 and DNMT3-R882 mutations, 99% samples for IDH1-R132 and IDH2-R172 mutations). HRM method reported no false-negative results, suggesting that it can be used for mutations screening. Moreover, HRM displayed much higher sensitivity in comparison with DNA sequencing in all assessed loci. With Sanger sequencing, robust calls were observed when the sample contained 50% of mutant DNA in the background of wild-type DNA. In marked contrast, the detection limit of HRM improved down to 10% of mutated DNA. Given the ubiquitous presence of wild-type DNA background in bone marrow aspirates and clonal variations regarding mutant allele burden, these results favor HRM as a sensitive, specific, labor-, and cost-effective tool for screening and detection of mutations in IDH1/2 and DNMT3A genes in patients with AML.

Acquired copy number changes are common in acute leukemia. They are reported as recurrent amplifications or deletions (del), and may be indicative of involvement of oncogenes or tumor suppressor genes in acquired disease, as well as serving as potential biomarkers for prognosis or as targets for molecular therapy. The present study reported a gain of copy number of 14q13 to 14q32, leading to immunoglobulin heavy chain locus splitting in a young adult female. To the best of our knowledge, this rearrangement has not been previously reported in B-cell acute lymphoblastic leukemia (ALL). Low resolution banding cytogenetics performed at the time of diagnosis revealed a normal karyotype. However, retrospective application of fluorescence in situ hybridization (FISH) banding and locus-specific FISH probes, as well as multiplex ligation-dependent probe amplification and high resolution array-comparative genomic hybridization, revealed previously hidden aberrations. Overall, a karyotype of 46, XX, del(9) (p21.3 p21.3),derivative(14) (pter-> q32.33:: q32.33-> q13 ::q32.33-> qter) was determined. The patient was treated according to the Polish Adult Leukemia Group protocol and achieved complete remission. The results of the present study indicate that a favorable prognosis is associated with these aberrations when the aforementioned treatment is administered.

Copy number variations (CNV) in CEBPA locus represent heterogeneous group of mutations accompanying acute myeloid leukemia (AML). The aim of this study was to characterize different CEBPA mutation categories in regard to biological data like age, cytology, CD7, and molecular markers, and identify possible factors affecting their etiology. We report here the incidence of 12.6% of CEBPA mutants in the population of 262 normal karyotype AML (NK-AML) patients. We confirmed that double mutant AMLs presented uniform biological features when compared to single CEBPA mutations and accompanied mostly younger patients. We hypothesized that pathogenesis of distinct CEBPA mutation categories might be influenced by different factors. The detailed sequence analysis revealed frequent breakpoint-associated microhomologies of 2 to 12bp. The analysis of distribution of microhomology motifs along CEBPA gene showed that longer stretches of microhomology at the mutational junctions were relatively rare by chance which suggests their functional role in the CEBPA mutagenesis. Additionally, accurate quantification of CEBPA transcript levels showed that double CEBPA mutations correlated with high-level CEBPA expression, whereas single N-terminal CEBPA mutations were associated with low-level CEBPA expression. This might suggest that high-level CEBPA expression and/or accessibility of CEBPA locus contribute to B-ZIP in-frame duplications.

Parvovirus B19 (B19V) is divided into three genotypes. Genotypes 2 and 3 may cause diagnostic difficulties and their epidemiology is not well understood. In the present study the prevalence of B19V genotypes in patients with symptomatic infection in Poland was evaluated and the course of infection in patients infected with non-genotype 1 strains is described. Real-time PCR, able to detect all three genotypes of B19V was used to screen patient plasma samples. Sixty-nine, mainly acute-phase B19V DNA positive cases were identified in patients from hematological and obstetric/gynecological wards between 2004 and 2008. Thirty patients were studied in greater detail and genotyping was performed by analysis of the NS1/VP1u region. The majority of samples were genotype 1. However two (6.6%) strains were identified as genotype 2, associated with high viremia and identified in a kidney transplant recipient with anemia and a leukemia patient, following chemotherapy, with pancytopenia. A change of immunosuppression treatment in the former and treatment with intravenous immunoglobulin in latter, resulted in normalization of clinical parameters, and whilst viral loads fell, B19V DNA was still detectable. The kidney transplant recipient subsequently became pregnant with no clinical complications, although persistently infected with B19V genotype 2. This is the first description of symptomatic cases of genotype 2 B19V infection in Eastern Europe suggesting that acute infection, particularly among immunocompromised patients with these virus strains may be more prevalent than thought.

The Guillain-Barre syndrome (GBS) could be a manifestation of neurotoxicity caused by multiple factors due to allogeneic bone marrow transplantations (alloBMT). In this paper we present a case of a 40-year old woman with chronic myeloid leukemia after alloBMT from her HLA identical brother. She developed the second grade of acute hepatic graft versus host disease (GvHD) shortly after alloBMT followed by chronic form. We observed Guillain-Barre syndrome probably triggering by GvHD in this patient.

Objectives:Cytogenetics is one of the most important prognostic factors in acute myeloid leukemia (AML). Two major, partially overlapping cytogenetic subsets of AML associated with an adverse prognosis, are AML with complex karyotype (CK) and AML with monosomal karyotype (MK). MK has been shown to constitute a cytogenetic feature that identifies AML patients (pts) with very poor outcomes when treated with currently available therapy. CK-AML pts are very heterogeneous cytogenetically, and treatment outcomes of these subsets are also influenced by TP53 alterations, complexity of the karyotype or presence of residual normal metaphases. However, impact of all these variables on outcome of CK-AML patients was so far analyzed separately. The aim of our study was to assess treatment outcome of CK-AML patients with or without MK regarding some cytogenetic and clinical features. Materials and methods: One hundred twenty five newly diagnosed AML patients with CK treated between January 2007 and January 2013 with PALG protocols (Holowiecki et al., JCO 2012) were included into the study. Cytogenetic analysis was performed on metaphases from bone marrow aspirates taken at diagnosis using standard banding techniques. Karyotypes were reported in accordance with the ISCN 2009. CK was defined as a presence of ≥ 3 unrelated abnormalities. MK was defined by the presence of an autosomal monosomy in conjunction with at least one other autosomal monosomy or structural abnormality in the absence of t(8;21), inv(16), t(15;17) (Breems et al., JCO 2008). FISH method was used to verify MK in all cases with monosomy as well as to assess 17p13 (TP53) deletion. Results:Clinical characteristics. The median age of all pts was 60 years (range 19-85 years). FISH verification proved the MK karyotype (MK+) in 75 out of 125 (60%) pts with CK-AML. Remaining pts had non-monosomal CK (MK-). Sixty six (55%) pts received intensive induction chemotherapy (IC) according to PALG protocols and 59 pts received non-intensive treatment with low-dose cytarabine (LDAC) or best supportive care (BSC). Baseline characteristics were generally balanced between MK+ and MK- groups in terms of age, WBC and PLT counts, hemoglobin level, percentage of bone marrow blasts at diagnosis and presence of residual normal metaphases in the karyotype. Higher proportions of MK+ pts had deletion of 17p13 (TP53) (47,9% vs. 20,4% of MK-; p=0.0017) as well as ≥5 cytogenetic aberrations (93,3% vs. 74%; p=0.004). Furthermore, there were no differences in the proportion of IC, LDAC and BSC strategies between both groups. Treatment Response and Outcome. With the median time of follow-up 7,7 months (mos), median OS in MK- pts was significantly longer compared to MK+ group (4,5 mos vs. 2,0 mos; p=0.0075). 66/125 CK-AML pts received standard IC according to PALG AML protocols (DA-36 pts; DAC-27 pts and DAF-3 pts). Only 7 pts underwent allogeneic stem cell transplantation (allo-SCT). In intensively treated pts the overall CR rate in MK- was about twice as high as in MK+ group (62,1% vs 32,4%; p=0.013). The median OS in MK- pts was longer compared to MK+ group (6,8 mos vs. 2,7 mos; p=0.016). Probability of 1-year OS in MK+ vs. MK- group was 14% vs. 30%. In multivariate analysis we have found that WBC >20 G/l, MK and ≥5 chromosomal abnormalities were independent prognostic factors associated with significantly shorter OS (Table 1). In contrast, allo-SCT was associated with survival benefit. Furthermore, we analyzed the impact of purine analogue containing induction regimens on treatment outcome. We found that addition of cladribine to the standard DA regimen improves OS in MK- but not in MK+ group (Figure 1). Conclusions: Monosomal karyotype, high WBC count and high complexity of the karyotype (≥5 aberrations) are associated with more aggressive clinical course and short survival in high-risk CK-AML. Cladribine added to standard DA induction regimen prolongs survival of pts with complex but not monosomal karyotype. *AW and EW equally contributed to the study. Figure 1 Figure 1. Figure 2 Figure 2. Disclosures No relevant conflicts of interest to declare.

Abstract 3537 Objectives: Cytogenetics is one of the most important prognostic factors in acute myeloid leukemia (AML). Breems et al., based on banding techniques (BT), have identified a monosomal karyotype (MK) to be associated with particularly poor survival. MK is defined by the presence of an autosomal monosomy in conjunction with at least one other autosomal monosomy or structural abnormality. However, classical BT may be not sufficient to confirm the “real” loss of a particular chromosome, especially in patients with complex karyotype (≥ 3 or ≥ 5 separate abnormalities) where parts of „missing” chromosomes could be involved in structural aberrations such as: additions, derivative chromosomes, rings, marker chromosomes and dicentrics. Molecular cytogenetic techniques, such as FISH, could be useful to verify if observed monosomy was “total” (loss of a complete chromosome) or “partial” (loss of only a part of chromosome including centromere). The aim of our study was to assess if the type of monosomy (total or partial), defined using FISH influences the prognosis of AML patients with MK. Materials and methods: Fifty newly diagnosed AML patients with MK, treated between January 2005 and January 2011 with PALG AML1/2004 and AML2/2004 protocols were included into the study. Cytogenetic analysis was performed on metaphases from bone marrow aspirates taken at diagnosis using standard BT. Karyotypes were centrally reviewed by two independent cytogeneticists and reported in accordance with the ISCN. In 41 cases FISH was performed with painting probes and in justified cases additionally with satellite probes and locus-specific probes to assess the type of monosomy. In 9 patients no confirmation by FISH was needed. Results: In 8/50 (16%) patients FISH verification proved the complex, but not monosomal (total or partial) karyotype. These patients were excluded from further study. The median age of 42 evaluable patients was 58 years (range 20–71 years). There were 26 males and 16 females. Twenty three (55%) patients received intensive induction chemotherapy according to PALG protocols (Holowiecki et al. Leukemia 2004), 8 patients - low dose cytarabine and 11 patients with high frailty index received treatment with hydroxyurea (n=9), and best supportive care (n=2). In 38/42 (90.5%) analyzed cases with MK abnormalities were complex. In all but one ≥5 separate aberrations were present. In 27/42 (64.3%) cases a total monosomy was confirmed by FISH (group A) and in 15/42 (35.7%) cases FISH revealed a monosomy to be the partial one (group B). Both groups A and B were comparable in terms of age, sex, immunophenotype and factors associated with tumor mass (leukemic bone marrow infiltration, WBC and peripheral blood blast count, as well as LDH activity). There was also no difference in the proportion of intensive and non-intensive treatment strategies between both groups. Six (26,1%) of intensively treated patients achieved complete remission (CR). The CR rate in patients with partial monosomy (40%) was higher than in total monosomy group (15,4%), however the difference was not significant (p=0.19; Fisher's exact test). The median overall survival (OS) for all evaluable patients was 66 days (range 1–659 days) and the probability of OS at 1 yr was 14% (95% CI 4–23%). Total monosomy was the only factor associated with decreased probability of OS both in univariate (p=0.036) (Fig. 1) and multivariate (p=0.037) analyses in AML patients with MK. Additionally, in patients with total monosomy the frequency of distinct monosomies was analyzed. The most frequent monosomies were monosomy 7 (n=13; 48%) and monosomy 18 (n=10; 37%). Other total monosomies included: monosomy 17 (n=3; 11%), monosomy 16 (n=2; 7%) and others (n=4). We did not observe any total monosomy 5. Conclusions: Our results indicate for the first time that partial monosomy is associated with significantly better OS than total monosomy in high risk AML with MK and provide further evidence for the heterogeneity of this defined cytogenetic group. These data also clearly demonstrate an important role of FISH method for precise evaluation of complex karyotype, which results in correct classification within MK group, as well as in adequate description of monosomy type. Disclosures: Wierzbowska: Genzyme: Membership on an entity's Board of Directors or advisory committees. Dmoszynska:Roche: Honoraria; Mundipharma:.

Abstract 1067 Elderly patients with acute myeloid leukemia are heterogenous group with poor outcome. All age, biological status and co-morbidities limit applicability of intensive chemotherapy. The PALG elaborated original system allowing stratification of patients aged >60 years to three groups with different therapeutic approach. Altogether 537 patients with newly diagnosed AML and median age 70 years (range 60–93) were classified as 1) ‘fit’ (n=163): age 60–79y, ECOG 0–2, proper liver and kidney function, without comorbidities, 2) ‘unfit’ (n=210): age >60 years, ECOG 0–2, normal liver and kidney function, comorbidities allowed, 3) ‘frail’ (n=164): ECOG 3–4. According to PALG 1/2005 protocol ‘fit’ patients were treated similarly as younger adults with daunorubicin (DNR, 3 days) + cytarabine (AraC, 7 days) +/− cladribine, followed by DNR + AraC consolidation and maintenance. ‘Unfit’ patients received either two courses of AraC+DNR (2+5) or AraC (5 days) + thioguanine + methotrexate, followed by manitenance. ‘Frail’ patients were considered for palliative cytoreduction and supportive care. Results: Complete remission (CR) rate was 35% for ‘fit’, 22% for ‘unfit’ and 0% for ‘frail’ patients. Median survival in the respective groups equaled 39 weeks, 26 w., and 14 w., while the probability of survival at 1 year was 39%, 27% and 10%. The rate of early (up to 8 weeks) mortality was 31%, 24% and 31%, respectively. In the Cox model the only factor independently affecting the risk of overall mortality in both ‘fit’ and x‘unfit’ group was serum LDH above upper quartile (HR=2, p=0.005 for ‘fit’, HR=1.65, p=0.006 for ‘unfit’). Among ‘frail’ patients the risk of mortality was increased in patients with performance status ECOG>2 (HR=1.85, p=0.0008), initial WBC >8.5×10e9/L (HR=1.65, p=0.006), and bone marrow blasts >58% (HR=1.8, p=0.001). We conclude that the proposed stratification system is feasible for elderly AML patients and represets a model for further developments of individualized therapeutic approaches. Survival of patients in whom remission induction therapy may be applied depends on initial tumor burden as reflected by high serum LDH level. The outcome of patients referred for palliative treatment depends additionally on initial performance status. In contrast, neither age nor karyotype were found to independently affect outcome in this study. Disclosures: No relevant conflicts of interest to declare.

Abstract 2138 In a previous study by the Polish Adult Leukemia Group (PALG 4–2002) we demonstrated that status of minimal residual disease (MRD) during induction-consolidation is the most important factor predicting long-term outcome of adult ALL (Br J Haematol 2008). In a new PALG 5–2007 protocol the treatment was intensified for patients with MRD level >0.1% of bone marrow cells and adjusted to age. Induction consisted of Epirubicine(4×), Vincristine(4×), Prednisone and PEG-asparaginase. Patients with complete remission (CR) but MRD >0.1% obtained additional 2nd phase of induction: AraC+Cyclophosphamide(Ctx) + 6MP. Consolidation consisted of 2 courses of Methotrexate(Mtx) + Etoposide + Dexamethasone and subsequent 2 cycles of Ctx + high dose AraC, asparaginase, intrathecal prophylaxis and CNS irradiation. Doses of Mtx were 500 mg/m2 for patients with MRD35y and 1500 mg/m2 for all remaining ones. For Ph+ ALL imatinib 600 mg/d was administered in parallel to chemotherapy. Patients with standard risk disease continued with 2 years maintenance while those with high-risk were referred for alloHSCT. High risk was defined as the presence of at least one of: initial WBC >30×10e9/L, age >35y, pro-B, early-T, mature-T, late CR, t(9;22), t(4;11) or MRD >0.1% at any time during induction-consolidation. Results of PALG 5–2007 protocol (N=108, median age 32y, range 16–55y) were compared to PALG 4–2002 (N=253, age 28y, range 16–55y), in which MRD status was not taken into account for treatment decisions. CR rate was 92% for PALG 5–2007 compared to 89% for PALG 4–2002. The probability of the overall survival at 36 months was 56% vs. 33% (p=0.02), while leukemia-free survival was 53% vs. 28% (p=0.04), respectively. The probability of relapse decreased from 54% in PALG 4–2002 to 16% in PALG 5–2007 (p=0.002). In a multivariate analysis adjusted to age, initial WBC and the presence of t(9;22) treatment according to PLAG 5–2007 protocol was associated with decreased risk of mortality (HR=0.57, p=0.02), relapse (HR=0.37, p=0.006) and treatment failure (HR=0.64, p=0.049). We conclude that individualized therapeutic approach with treatment intensity adjusted to MRD status and age may result in improved outcome of adults with ALL. *This multi-institutional study was supported by Polish Ministry of Sciences Grant NN-402366433 Disclosures: Off Label Use: Dasatinib as first line therapy in Ph ALL.

Abstract 2055 Poster Board II-32 This trial is a continuation of earlier Polish Adult Leukemia Group (PALG) studies on the use of purine analogues for therapy of AML patients (Leukemia 2004,18:989–97, updated at 7 y: ASH 2006, Abstr.N#2003, Ann Hematol. 2008, 87:361–7). The goal of the present study was to evaluate the efficacy of combination including fludarabine – DAF (daunorubicine 60 mg/m2/d iv, d 1–3; cytarabine (AraC) 200 mg/m2/d ci, d 1–7, and fludarabine 25 mg/m2 iv d 1–5) in untreated adult patients with AML, based on head to head comparison with DAC (DNR, AraC, Cladribine), and standard DA regimens (preliminary results: ASH 2008, abstr.#133). Primary end-points were: complete remission rate (CR) and overall survival (OS); secondary objectives were: toxicity and leukemia-free survival (LFS). Patients achieving CR received two courses of subsequent intensive consolidation: HAM (HD AraC, mitoxantrone) and HD AraC. Reduction of consolidation was accepted in patients treated with early alloBMT. In case of partial remission (PR) after the first induction course the same regimen was repeated. Patients with no remission (NR) or with PR after 2 induction courses were withdrawn from the study. Between 09.2004 and 05.2008, 673 adult untreated AML patients aged 18–60 y, median 47 y, sex: male 49,5%, female 50,5%, treated in 18 co-operating PALG centers were centrally randomized to either DAF (n=225), DAC (n=224) or DA (n=224) arm (1:1:1). PML/RAR alfa positive - FAB M3 cases were excluded. The study groups were well balanced in respect of age, sex, FAB subtype, and WBC. The results are summarized in the table. Outcome DAC (n = 223) DAF (n = 219) DA (n = 210) P Value (DAC vs DA) P Value (DAC vs DAF) CR 68 59 56 .013 .08 CR after 1 cycle 62 55 50,5 .017 16 3-yr OS 46 30 31 02 .02 2-yr LFS 47 40 39 NS NS Both, the entire CR rate and the CR rate after a single induction course were significantly superior in the DAC arm if compared with DA and DAF subgroups. With a median follow-up of 34 months (the longest observation time 5y) the OS rate equaled 46% for the DAC treated subgroup and was higher in comparison to the standard DA arm and the DAF arm. There were no significant differences in the leukemia free survival rates. The early death rates of 8,5–11%. were similar in the studied treatment subgroups. All patients developed WHO grade IV thrombocytopenia and agranulocytosis. The frequency and severity of infections, mucositis, vomiting, diarrhoea, alopecia, polyneuropathy as well as of cardiac, liver or kidney dysfunctions were comparable in particular arms. In conclusion, this updated results of randomized study prove that the incorporation of cladribine to the standard DA induction regimen (DAC) improves CR rate and the overall survival in adults with AML aged up to 60 y, without additional toxicity. This beneficial effect was not observed in patients treated using the DAF protocol with fludarabine added to the standard “DA 3+7” schedule. Disclosures: Robak: Celgene: Consultancy; Roche: Honoraria, Research Funding; Genmab: Research Funding; Cambridge Antibody Technology: Research Funding; GlaxoSmithKline: Honoraria. Warzocha:BMS: Consultancy, Honoraria; Celgene: Consultancy; Roche: Honoraria; Pfizer: Honoraria; Amgen: Honoraria.

Our previous PALG study in 445 de novo AML patients has demonstrated that addition of cladribine to the standard daunorubicine-cytarabine (DA) remission induction regimen - DAC has a beneficial influence on both the CR rate after one induction cycle (p=0,0008) and on survival in patients older than 40 y (Leukemia2004,18:989–97, updated at 7 y: ASH 2006, Abstr.N#2003). The goal of this study was to evaluate the efficacy of original combination including another purine analogue fludarabine – DAF (daunorubicine 60 mg/m2/d iv, d 1–3; cytarabine (AraC) 200 mg/m2/d ci, d 1–7, and fludarabine 25 mg/m2 iv d 1–5) in untreated patients with AML, aged up to 60 y, based on head to head comparison with DAC - (DNR, AraC, Cladribine), and standard DA regimens. We evaluated earlier the DAF protocol in relapsed or refractory AML (PALG pilot study; Ann Hematol.2008, 87:361–7. Epub 2007 Dec 12); the tolerance was good, CR 44%, LFS 38%. Primary objectives of the presented trial were: complete remission rate (CR) and overall survival (OS); secondary objectives were: toxicity and leukemia-free survival (LFS). Additional analysis was planned for patients submitted to an early bone marrow allotransplantation (alloBMT) after CR. Patients achieving CR received two courses of subsequent intensive consolidation: HAM (HD AraC, mitoxantrone) HD AraC. Reduction of consolidation was accepted in patients treated with early alloBMT. In case of partial remission (PR) after the first induction course the same regimen was repeated. Patients with no remission (NR) or with PR after 2 induction courses were withdrawn from the study. Between 09.2004 and 05.2008, 673 adult untreated AML patients aged 18–60 y, median 47 y, sex: male 49,5%, female 50,5%, treated in 18 co-operating Polish Adult leukemia Group (PALG) centres were centrally randomised to either DAF (n=225), DAC (n=224) or DA (n=224) arm (1:1:1). PML/RAR alfa positive - FAB M3 cases were excluded. The study groups were well balanced in respect of age, sex, FAB subtype, and WBC. After a single induction course the CR rate for patients receiving DAC equalling 63% was significantly higher if compared with DA - 51% (p=0,01), whereas no significant differences were noted between DAC vs. DAF - 55% and DAF vs. DA subgroups. Also the entire CR rate of 68% in the DAC arm was higher in comparison with DA one - 57% (p=0,02). No significant differences were found between DAC vs. DAF - 60%, and DAF vs. DA. At median time of 24 months (longest observation time 3,5y) the OS rate equalled 51% for the DAC treated subgroup and was higher in comparison to the standard DA arm - 39% and the DAF arm - 36% (p=0,03). The leukemia free survival rates (LFS) in DAC, DA and DAF treated cohorts equalled 51%, 32% and 41% respectively (p=NS). The early death rates of 8–10%. were similar in the studied treatment subgroups. All patients developed WHO grade IV thrombocytopenia and agranulocytosis. The frequency and severity of infections, mucositis, vomiting, diarrhoea, alopecia, polyneuropathy as well as of cardiac, liver or kidney dysfunctions were comparable in particular arms. In conclusion, this randomised study proves that the addition of cladribine to the standard DA induction regimen (DAC) improves CR rate and the overall survival in adults with AML aged up to 60 y, without additional toxicity. This beneficial effect was not observed in patients treated using the DAF protocol with fludarabine added to the standard “DA 3+7” schedule

Chromosomal aberrations analyzed by cytogenetic and molecular methods are major prognostic factors determining treatment options in patients with acute leukemias. The aim of this study was to compare cytogenetic and molecular methods as diagnostic and prognostic tools in patients with acute leukemias. Sixty one previously untreated patients with acute leukemias (AML - 43, ALL - 18) were studied for the presence of chromosomal translocations and corresponding fusion genes [AML1-ETO t(8;21), CBFB-MYH11 inv(16), PML-RARA t(15;17), BCR-ABL p190, p210 t(9;22), MLL-AF4 t(4;11), TEL-AML1 t(12;21), E2A-PBX1 t(1;19), SIL-TAL1 del(1)] using both molecular (RT-PCR, nested PCR) and cytogenetic (GTG) methods. Molecular diagnostics was performed from RNA isolated from bone marrow samples according to BIOMED-1 protocol. Cytogenetic studies were carried out with classical GTG and FISH methods. G-banded mitoses of bone marrow specimen were analysed according to ISCN. Chromosomal aberrations were found in 32.8% patients using GTG method while the parallel molecular tests revealed related fusion genes in 50.8% patients. In 8.5% patients cytogenetic analysis was not performed because of lack of metaphases in cultured cells. All cytogenetic aberrations found in GTG were also confirmed in RT-PCR. Stratification into cytogenetic risk groups was performed after applying combined analysis of karyotyping and molecular tests. Low cytogenetic risk group consisted of 32.8% patients including 11.5% diagnosed with GTG and additionally 21.3% patients after applying molecular tests. The intermediate and high cytogenetic risk group consisted of 32.8% and 34.4% respectively using combined cytogenetic and molecular diagnostics. In low cytogenetic risk group, 85% of patients achieved complete remission (CR), early deaths were found in 15% and none of the patients presented primary chemotherapy resistance. In intermediate risk group CR were obtained in 80%, chemoresistance in 10% and early deaths were observed in 10% of patients. In high cytogenetic risk group, CR were achieved in only 23.8% and chemoresistance occured in 76.2% of the patients. In conclusion we suggest that molecular and cytogenetic tests are complementary methods and should be used in parallel in the initial diagnosis of patients with acute leukemias. This seems to be critical for obtaining the accurate diagnosis, cytogenetic risk assessment and choosing an optimal treatment options.